Cellulase Immobilization onto Magnetic Halloysite Nanotubes: Enhanced Enzyme Activity and Stability with High Cellulose Saccharification

Abstract

A quest for efficient biotransformation of cellulosic material into sustainable biochemical products for recent biotechnological interventions is currently under way. Herein, we report the fabrication of nanobiocatalyst (NBC) employing halloysite nanotubes (HNTs) as a template for immobilizing cellulase enzyme, which catalyzed the hydrolysis of cellulose into glucose. Magnetic character was imported to HNTs by in situ anchoring of iron oxide nanoparticles, onto which cellulase was immobilized using aminosilane surface-functional chemistry. Characterization studies revealed nanobiocatalyst to be extremely stable during heterogeneous catalysis without compromising their catalytic activity. The optimization of process parameters yielded ∼93.5% activity of cellulase with high enzyme loading (111.6 mg·g–1 HNTs) after immobilization. Immobilized cellulase displayed superior stability at elevated temperatures (≥60°C) and storage capability compared with their free forms. The NBC even retained ∼68.2% of its original activity after seven consecutive uses with a minimum yield of 25.4 mg glucose·g–1 cellulose and was 100% recoverable using a magnet. Displaying a high ionic-liquid tolerance ability is concurrent with superior catalytic potential against CMC and extracted cellulose (bagasse), and achieving ∼50.2% saccharification and 0.56 g glucose·g–1 cellulose within 48 h of continuous operation establishes the commercial viability of using cellulase-immobilized HNTs for efficient cellulose hydrolysis. The sustainability and eco-friendly endeavors in this approach would pave the way toward valorization and consolidated bioprocessing of cellulose materials.

Bioreactor and Bioprocess Design Issues in Enzymatic Hydrolysis of Lignocellulosic Biomass

“…More sophisticated approaches to enzyme retention have been investigated by immobilizing the enzyme on specific supports such as nanofibers, nanoparticles and organic metal frames. Sillu et al [64] proposed the immobilization of cellulase on magnetic halloysite nanotubes. Driven by the potential ease of the recovery, the use of magnetic CLEA (Cross Linked Enzyme Aggregates) of cellulase for the hydrolysis of cellulose has been successfully reported at lab scale by several contributions [65][66][67].…”

Magnetic Nanomaterials as Biocatalyst Carriers for Biomass Processing: Immobilization Strategies, Reusability, and Applications

“…Sillu & Agnihotri [37] developed magnetic halloysite nanotubes (MHNTs), a different conformation consisting of silica-based nanotubes of Al 2 SiO 5 (OH) 4 .2H 2 O that present silanol (-Si-OH) and siloxane groups (-Si-O-Si-) at the surface. The first step was the coating of MNPs in situ with halloysite nanotubes; then, it was grafted with amino groups using 3-APTES and activated, resulting in protein loading of 80.48% and cellulase activity yield of about 93.5%.…”

“…It is essential to note that the optimization step is fundamental for immobilized enzyme efficiency, catalytic activity, stability, and recycling ability. The studies that previously determined the optimal immobilization parameters have usually shown better yields and efficiencies in enzyme immobilization [12,37,47,52,56,64,65].…”

Past, Present and Future Perspectives on Halloysite Clay Minerals

“…Similarly, in 2020 cellulase enzyme was grafted on magnetic HNTs’ (MHNTs’) external surface for biocatalytic purposes ( Figure 9 ) [ 121 ]. It was demonstrated that the nanomaterial exhibited an excellent catalytic activity at elevated temperatures, ionic liquid-tolerant characteristics, hydrolyzation continuously over longer durations, and eco-friendly attributes, features which made the cellulase-MHNTs amenable for high cellulose conversion.…”

“… Schematic representation of the synthesis of cellulase-MHNTs (magnetic HNTs). Reproduced with permission from [ 121 ]. …”